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PCR-RFLP typification of microbes used in the production of a fermented fish product

Spengler, C. J. (2001-12)

Thesis (MScFoodSc)--Stellenbosch University, 2001.

Thesis

ENGLISH ABSTRACT: The preservation of various fresh fish products is achieved by either smoking,
salting, canning, freezing or fermenting a highly perishable raw product. Since
many of these facilities are not readily available, the use of fermentation as a
means of preserving the product has been extensively practiced. However, the
fermentation of fish is a time consuming practise and only by accelerating the
process would it be possible to ensure the production of a more cost effective and
readily available safe end-product.
The quality of the fermented fish product is partially determined by the
fermentation conditions and the metabolic activity of the microbes present. The
rapid identification of the microbes present during the fermentation would enable
the selection of possible starters to ensure an accelerated production of high
quality fermented fish products. This study was thus undertaken to develop
identification fingerprints for bacteria isolated from fermented fish products. A
1300 bp fragment of the 16S rRNA genes of each of the bacteria previously
isolated was successfully amplified using the PCR technique. The isolates
included strains of the genera Bacillus, Staphylococcus, Sphingomonas, Kocuria,
Brevibacillus, Cryseomonas, Vibrio, Stenotrophomonas and Agrobacterium. The
data obtained can, therefore, be used in the identification of these microbes
isolated from other similar fermented fish products. The fingerprints could also be
used to assist in determining the dominant microbial populations responsible for
the characteristic qualitative changes occurring in the fish product during
fermentation.
The microbial composition of a fermenting fish product partially determines
the quality of the end-product, therefore, the use of selected bacterial starters
could result in the accelerated production of a microbial safe fermented fish
product. A further objective of this study was to accelerate the production of a
fermented fish product by inoculating macerated trout with either selected lactic
acid bacteria (LAB) or with selected bacteria with high proteolytic activity over a 30
day fermentation period. The LAB included a combination of Lactobacillus
plantarum, Lactococcus diacetylactis and Pediococcus cerevisiae strains, whereas
the bacteria with high proteolytic activity included strains of Kocuria varians,
Bacillus subtilis, two strains of B. amyloliquefaciens and a combination of these bacterial species. The quality of the fermented product was determined using
changes in product pH, titratable acidity (%TA) and free amino nitrogen (FAN)
formation as efficiency parameters.
The data obtained during the fermentation of the macerated trout showed
that the selected starters did not have a significant effect on the pH decrease in
the product over a 30 day fermentation period. The LAB strains did not have a
significant effect on the %TA of the fermenting fish product, yet the presence of
these bacteria appeared to limit the FAN production in the product. The bacteria
with high proteolytic activity resulted in slightly enhanced %TA values and a higher
FAN content in the fermented product. It was also determined that the LAB and
Kocuria varians, in contrast to the Bacillus spp. inoculums, did not survive the
fermentation conditions well, possibly due to the low pH environment. The
presence of the starter bacteria in the fermenting fish mixture at the end of the
fermentation was also successfully determined with the use of the PCR-RFLP
technique.
The fermented fish product, obtained at the end of the fermentation period,
had a good aroma and compared favourably to similar commercially available
fermented fish products. The use of different microbial starters could in future
enable the production of a diverse range of high quality products, which could be
produced and marketed locally.